Difference between revisions of "Team:Aix-Marseille/Notebook"
(→Thursday, September 8th) |
m |
||
| (46 intermediate revisions by 2 users not shown) | |||
| Line 1: | Line 1: | ||
| − | |||
__NOTOC__ | __NOTOC__ | ||
| − | + | {{:Team:Aix-Marseille/Template-Style-and-Menu}}{{:Team:Aix-Marseille/Template-Notebook}}<div class="events"> | |
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | + | ||
| − | <div class="events"> | + | |
| − | + | ||
<div id="2016-06-13" class="evday"> | <div id="2016-06-13" class="evday"> | ||
| − | + | === Monday, June 13th === | |
* Preparation of heat competent DH5α cells ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol #1]). | * Preparation of heat competent DH5α cells ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol #1]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-14" class="evday"> | <div id="2016-06-14" class="evday"> | ||
| − | + | === Tuesday, June 14th === | |
* Transformation of DH5α cells with test plasmid ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of DH5α cells with test plasmid ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-15" class="evday"> | <div id="2016-06-15" class="evday"> | ||
| − | + | === Wednesday, June 15th === | |
* Preparation of heat competent Tg1 cells ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). | * Preparation of heat competent Tg1 cells ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). | ||
* Transformation of Tg1 cells with test plasmid ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of Tg1 cells with test plasmid ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-06-16" class="evday"> | <div id="2016-06-16" class="evday"> | ||
| − | + | === Thursday, June 16th === | |
* Transformation of DH5α cells with pSB1C3 + RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of DH5α cells with pSB1C3 + RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Transformation with K880005, K823017, I0500, K1404006, B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation with K880005, K823017, I0500, K1404006, B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-17" class="evday"> | <div id="2016-06-17" class="evday"> | ||
| − | + | === Friday, June 17th === | |
* Plasmids purification K880005, K823017, I0500, K1404006, B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmids purification K880005, K823017, I0500, K1404006, B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Plasmids purification pSB1C3 + RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmids purification pSB1C3 + RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-20" class="evday"> | <div id="2016-06-20" class="evday"> | ||
| − | + | === Monday, June 20th === | |
* pSB1C3 digestion from 17/06 plasmids purification ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Digestion_protocol_BioBrick_Assembly_Kit Protocol#4]). | * pSB1C3 digestion from 17/06 plasmids purification ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Digestion_protocol_BioBrick_Assembly_Kit Protocol#4]). | ||
| Line 180: | Line 42: | ||
* Biobricks plasmids purifications gel migration ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Biobricks plasmids purifications gel migration ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Transformation Tg1 with RFP+pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]) | * Transformation Tg1 with RFP+pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]) | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-21" class="evday"> | <div id="2016-06-21" class="evday"> | ||
| − | + | === Tuesday, June 21th === | |
* pSB1C3 digestion and gel migration, NucleoSpin gel extract and PCR cleanup ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Digestion_protocol_BioBrick_Assembly_Kit Protocol#4]). | * pSB1C3 digestion and gel migration, NucleoSpin gel extract and PCR cleanup ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Digestion_protocol_BioBrick_Assembly_Kit Protocol#4]). | ||
* E. Coli and Desulfo FliC (E/P digested) ligation in linearized pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * E. Coli and Desulfo FliC (E/P digested) ligation in linearized pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Transformation of pSB1C3 RFP in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of pSB1C3 RFP in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-22" class="evday"> | <div id="2016-06-22" class="evday"> | ||
| − | + | === Wednesday, June 22th === | |
* Transformation in DH5α of ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]): | * Transformation in DH5α of ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]): | ||
| Line 205: | Line 63: | ||
* Plasmid purification B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Transformation in DH5α of interlab studies plasmids ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation in DH5α of interlab studies plasmids ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-23" class="evday"> | <div id="2016-06-23" class="evday"> | ||
| − | + | === Thursday, June 23th === | |
| − | + | [[File:T--Aix-Marseille--PCR-23-06.jpg|left|thumb|<u>Electrophoresis revelation</u> of E/P digestion on pSB1C3. Awaited size of 2070 bp. The two samples are the same digestion and we did a gel purification on the awaited band.]] | |
* Digestion E/P pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion E/P pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Purification pSB1C3 purified linearized 23/06 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.236_:_Digestion_and_ligation_for_verification_and_BioBrick_Assembly Protocol#6]). | * Purification pSB1C3 purified linearized 23/06 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.236_:_Digestion_and_ligation_for_verification_and_BioBrick_Assembly Protocol#6]). | ||
| Line 219: | Line 75: | ||
* Transformation in Tg1 and DH5α ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation in Tg1 and DH5α ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-06-24" class="evday"> | <div id="2016-06-24" class="evday"> | ||
| − | + | === Friday, June 24th === | |
| − | + | [[File:T--Aix-Marseille--PCR-24-06.jpg|left|thumb|Verification of the plasmid purifications on pSB1C3 pSB1CA3 pSB1CK3 pSB1CT3 (the name are wroten on the top of the gel) the awaited sizes are 2070bp.]] | |
* Digestion E/P pSB1C3, pSB1A3, pSB1K3, pSB1T3, and agarose gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion E/P pSB1C3, pSB1A3, pSB1K3, pSB1T3, and agarose gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Colony PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]) of | * Colony PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]) of | ||
| Line 236: | Line 91: | ||
* Redone transformation test device 1 and 3 interlab studies ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Redone transformation test device 1 and 3 interlab studies ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Plasmid purification pSB1C3 pSB1CA3 pSB1CK3 pSB1CT3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification pSB1C3 pSB1CA3 pSB1CK3 pSB1CT3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| − | |||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-27" class="evday"> | <div id="2016-06-27" class="evday"> | ||
| − | + | === Monday, June 27th === | |
* Transformations DesA, DesB, DesC, DesD, FliC E.Coli and desulfo, interlab studies ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformations DesA, DesB, DesC, DesD, FliC E.Coli and desulfo, interlab studies ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Transformations failed ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformations failed ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Transformation pSB1T3 plasmid purification, pSB1K3 plasmid purification, control ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.234_:_Plasmid_DNA_purification Protocol#4] and[https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation pSB1T3 plasmid purification, pSB1K3 plasmid purification, control ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.234_:_Plasmid_DNA_purification Protocol#4] and[https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-28" class="evday"> | <div id="2016-06-28" class="evday"> | ||
| − | + | === Tuesday, June 28th === | |
* Plasmid purification promoter+RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification promoter+RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* PCR of IDT sequences (fliC and DesA to D) and gel check ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR of IDT sequences (fliC and DesA to D) and gel check ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Gel extract pSB1CK3, pSB1CC3, pSB1CT3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * Gel extract pSB1CK3, pSB1CC3, pSB1CT3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-06-29" class="evday"> | <div id="2016-06-29" class="evday"> | ||
| − | + | === Wednesday, June 29th === | |
* Plasmid purification from interlab studies, T3 and K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification from interlab studies, T3 and K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| Line 268: | Line 116: | ||
* Redone PCR of IDT sequences (fliC and DesA to D) (different Tm) and gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * Redone PCR of IDT sequences (fliC and DesA to D) (different Tm) and gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
* Transformation of fliCs from remaining ligation in Tg1 Sandra ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of fliCs from remaining ligation in Tg1 Sandra ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-06-30" class="evday"> | <div id="2016-06-30" class="evday"> | ||
| − | + | === Thursday, June 30th === | |
* Migration FliC coli, desulfo, DesB, DesC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Migration FliC coli, desulfo, DesB, DesC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-01" class="evday"> | <div id="2016-07-01" class="evday"> | ||
| − | + | === Friday, July 1st === | |
| − | + | [[File:T--Aix-Marseille--PCR-29-06.jpg|left|thumb|Verification of the plasmid purifications on pSB1C3.]] | |
* Digestion PCR products and vector pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion PCR products and vector pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* PCR clean up on digested pSB1C3, fliC coli desulfo, DesB and DesC, migration and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up on digested pSB1C3, fliC coli desulfo, DesB and DesC, migration and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | |||
| − | |||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-04" class="evday"> | <div id="2016-07-04" class="evday"> | ||
| − | + | === Monday, July 4th === | |
* Isolated white colonies on dishes | * Isolated white colonies on dishes | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-05" class="evday"> | <div id="2016-07-05" class="evday"> | ||
| − | + | === Tuesday, July 5th === | |
* Reisolated colonies from 04/07 | * Reisolated colonies from 04/07 | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-06" class="evday"> | <div id="2016-07-06" class="evday"> | ||
| − | + | === Wednesday, July 6th === | |
* Colony PCR with FLiC E.Coli Desulfo DesB DesC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR with FLiC E.Coli Desulfo DesB DesC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-07" class="evday"> | <div id="2016-07-07" class="evday"> | ||
| − | + | === Thursday, July 7th === | |
| − | + | [[File:T--Aix-Marseille--PCR-07-07.jpg|left|thumb|Colony PCR from 06/07. For the two first holes, the awaited size is 1547 bp, for the two last, this size is 994bp]] | |
* Plasmid purification DesC DesB FliC coli ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification DesC DesB FliC coli ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* PCR on A2 A4 B1 clones from 6/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR on A2 A4 B1 clones from 6/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Digestion E/P of A5 B4 C3 C2 minipreps and pSB1A3, pSB1C3, pSB1K3, pSB1T3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion E/P of A5 B4 C3 C2 minipreps and pSB1A3, pSB1C3, pSB1K3, pSB1T3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | |||
| − | |||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-08" class="evday"> | <div id="2016-07-08" class="evday"> | ||
| − | + | === Friday, July 8th === | |
* Transformation BOO34 and I0500 in pSB1K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation BOO34 and I0500 in pSB1K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| Line 335: | Line 165: | ||
* PCR clean up of vectors ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up of vectors ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
* PCR SLIC FliC coli/desulfo, DesA, B, C, D, csgA -> failed ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR SLIC FliC coli/desulfo, DesA, B, C, D, csgA -> failed ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-11" class="evday"> | <div id="2016-07-11" class="evday"> | ||
| − | + | === Monday, July 11th === | |
* Redone PCR SLIC 8/8 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | * Redone PCR SLIC 8/8 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | ||
| Line 347: | Line 175: | ||
* Transformation Tg1 with fliC coli desulfo, desA, desB in pSB1A3 from SLIC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation Tg1 with fliC coli desulfo, desA, desB in pSB1A3 from SLIC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* PCR clean up desA, desB, fliC coli desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up desA, desB, fliC coli desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-12" class="evday"> | <div id="2016-07-12" class="evday"> | ||
| − | + | === Tuesday, July 12th === | |
* Purification clean up DesC and CsgA from PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * Purification clean up DesC and CsgA from PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| Line 360: | Line 186: | ||
* Transformation SLIC mix in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation SLIC mix in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Transformation constructions #7 (kanamycin) 9 and 55 (chloramphenicol) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation constructions #7 (kanamycin) 9 and 55 (chloramphenicol) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-13" class="evday"> | <div id="2016-07-13" class="evday"> | ||
| − | + | === Wednesday, July 13th === | |
* Plasmid purification DesA, DesB, fliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification DesA, DesB, fliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| Line 371: | Line 195: | ||
* Colony PCR #1 2 3 50 51 53 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR #1 2 3 50 51 53 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Plasmid purification pSB1 A/C/K/T 3, gel migration ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification pSB1 A/C/K/T 3, gel migration ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-18" class="evday"> | <div id="2016-07-18" class="evday"> | ||
| − | + | === Monday, July 18th === | |
* Ligation (3A) in pSB1C3 of desA+RBS, desB+RBS, fliC desulfo+RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation (3A) in pSB1C3 of desA+RBS, desB+RBS, fliC desulfo+RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| Line 384: | Line 206: | ||
* Digestion E/P and PCR clean up empty vectors A3 C3 K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion E/P and PCR clean up empty vectors A3 C3 K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* PCR clean up on A3, C3 and K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up on A3, C3 and K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-19" class="evday"> | <div id="2016-07-19" class="evday"> | ||
| − | + | === Tuesday, July 19th === | |
* Colony PCR on DesA-RBS, DesB-RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR on DesA-RBS, DesB-RBS ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Ligation and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Ligation and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* Interlab studies: transformation in DH5α ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Interlab studies: transformation in DH5α ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-20" class="evday"> | <div id="2016-07-20" class="evday"> | ||
| − | + | === Wednesday, July 20th === | |
* Colony PCR on DesA-RFP, DesC-RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR on DesA-RFP, DesC-RFP ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Plasmid purification gel migration DesB rbs, DesA rbs, DesB ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification gel migration DesB rbs, DesA rbs, DesB ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-21" class="evday"> | <div id="2016-07-21" class="evday"> | ||
| − | + | === Thursday, July 21th === | |
* Interlab studies: measurements plate reader | * Interlab studies: measurements plate reader | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-22" class="evday"> | <div id="2016-07-22" class="evday"> | ||
| − | + | === Friday, July 22th === | |
* Colony PCR on FliC, FliD, DesA DesB and CsgA ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR on FliC, FliD, DesA DesB and CsgA ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-25" class="evday"> | <div id="2016-07-25" class="evday"> | ||
| − | + | === Monday, July 25th === | |
* Redone 22/7 Colony PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Redone 22/7 Colony PCR ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Plasmid purification DesC Csg1 DesC+RBS and digestion E/P ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification DesC Csg1 DesC+RBS and digestion E/P ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Digestion E/P DesA+RBS, DesB ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion E/P DesA+RBS, DesB ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-26" class="evday"> | <div id="2016-07-26" class="evday"> | ||
| − | + | === Tuesday, July 26th === | |
* Competent Tg1 preparation ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). | * Competent Tg1 preparation ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-27" class="evday"> | <div id="2016-07-27" class="evday"> | ||
| − | + | === Wednesday, July 27th === | |
* Digestion K88005 E/S, fliC desulfo X/P, pSB1K3 E/P ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion K88005 E/S, fliC desulfo X/P, pSB1K3 E/P ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-28" class="evday"> | <div id="2016-07-28" class="evday"> | ||
| − | + | === Thursday, July 28th === | |
* Colony PCR from respread dishes 27/7 on DesC-RBS, DesA-7.8, DesA-7.1, FliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR from respread dishes 27/7 on DesC-RBS, DesA-7.8, DesA-7.1, FliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* PCR gradient on csgA (BBa140034) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR gradient on csgA (BBa140034) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-07-29" class="evday"> | <div id="2016-07-29" class="evday"> | ||
| − | + | === Friday, July 29th === | |
* Plasmid purification DesA DesC, E/P digestion ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Plasmid purification DesA DesC, E/P digestion ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation 3A pSB1K3 RBS FliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation 3A pSB1K3 RBS FliC desulfo ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Redone gel migration PCR 28/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Redone gel migration PCR 28/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-08-01" class="evday"> | <div id="2016-08-01" class="evday"> | ||
| − | + | === Monday, August 1st === | |
* Transformation ligation 29/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation ligation 29/7 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
* PCR amplification IDT sequences DesA/B/C fliC coli and oligo, gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR amplification IDT sequences DesA/B/C fliC coli and oligo, gel extract ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3] and [https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-08-02" class="evday"> | <div id="2016-08-02" class="evday"> | ||
| − | + | === Tuesday, August 2rd === | |
| − | + | [[File:T--Aix-Marseille--PCR-02-08.jpg|left|thumb|Digestion of backbones pSB1C3, pSB1A3 and pSB1K3 verifications. Like all our backbone digestions, the awaited size is 1070bp.]] | |
* Colony PCR of CsgA from the 01/08 using EconoTac ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR of CsgA from the 01/08 using EconoTac ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* PCR clean up of DesC(200ng/µL) and gel extraction of DesA (83ng/µL), DesB(53ng/µL), FliC <i>E. coli</i> ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up of DesC(200ng/µL) and gel extraction of DesA (83ng/µL), DesB(53ng/µL), FliC <i>E. coli</i> ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | * SLIC of DesA, DesB, DesC, DesD and transformation of 10µL in 100µL | + | * SLIC of DesA, DesB, DesC, DesD and transformation of 10µL in 100µL competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). |
* Sequencage of Desulfovribrio clone 3 validated. | * Sequencage of Desulfovribrio clone 3 validated. | ||
* E/P digestion on pSB1C3, pSB1A3 and pSB1K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * E/P digestion on pSB1C3, pSB1A3 and pSB1K3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.235_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | |||
| − | |||
| − | |||
</div><div id="2016-08-03" class="evday"> | </div><div id="2016-08-03" class="evday"> | ||
| − | + | === Wednesday, August 3th === | |
* Digestion Bba_B0034 by EcoRI/XbaI (pSB1A3) and Bba_I0500 by EcoRI/(pSB1C3) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion Bba_B0034 by EcoRI/XbaI (pSB1A3) and Bba_I0500 by EcoRI/(pSB1C3) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| Line 497: | Line 294: | ||
- by 3A technical : Bba_K1951005 and Bba_K880005 digested the 07.18.2016 | - by 3A technical : Bba_K1951005 and Bba_K880005 digested the 07.18.2016 | ||
* Transformation of BBa_B0034+BBa_I0500 ligated in Tg1 and stread on Amp LB agar petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of BBa_B0034+BBa_I0500 ligated in Tg1 and stread on Amp LB agar petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| + | </div><div id="2016-08-04" class="evday"> | ||
| − | + | === Thursday, August 4th === | |
| − | + | ||
| − | + | ||
| − | + | ||
* Transformation of Bba_K1951008 (FliC <i>E. coli</i> and spread on kanamycin LB agar petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of Bba_K1951008 (FliC <i>E. coli</i> and spread on kanamycin LB agar petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| Line 507: | Line 302: | ||
* Colony PCR from the transformed bacteria of 08.03.2016 and starter of the good clones have been sent ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR from the transformed bacteria of 08.03.2016 and starter of the good clones have been sent ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Plasmid purification of DesA, clone3(141.47ng/µL), FliC <i>E. coli </i> clone 2 (169,76ng/µL) and FliC Desulfo (133,47ng/µL) valided to the sequencage have been made ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification of DesA, clone3(141.47ng/µL), FliC <i>E. coli </i> clone 2 (169,76ng/µL) and FliC Desulfo (133,47ng/µL) valided to the sequencage have been made ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| − | + | </div> | |
| − | + | ||
| − | + | ||
<div id="2016-08-05" class="evday"> | <div id="2016-08-05" class="evday"> | ||
| − | + | === Friday, August 5th === | |
* Colony PCR from the transformed bacteria of 08.03.2016 and starter of the good clones have been sent ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR from the transformed bacteria of 08.03.2016 and starter of the good clones have been sent ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* SLIC of CsgA in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | * SLIC of CsgA in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | ||
* Sent to the sequencing Bba_K1951009 and BBa_B0034+BBa_I0500. | * Sent to the sequencing Bba_K1951009 and BBa_B0034+BBa_I0500. | ||
| − | + | </div> | |
| − | + | ||
<div id="2016-08-08" class="evday"> | <div id="2016-08-08" class="evday"> | ||
| − | + | === Monday, August 8th === | |
| − | + | [[File:T--Aix-Marseille--PCR-08-08.jpg|left|thumb|Colony PCR verification. The clones picked up on the plates are wroten on the right of the gel. the awaited size is 506bp for all clones.]] | |
* Ligation of BBa_B0034+BBa_I0500 and Bba_K1951000 into pSB1K3 to create desA producer ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of BBa_B0034+BBa_I0500 and Bba_K1951000 into pSB1K3 to create desA producer ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Transformation of the previous ligation in | + | * Transformation of the previous ligation in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations protocol#2]). |
* Starters from DesC and Bba_K1951008 of the 08.04.2016. | * Starters from DesC and Bba_K1951008 of the 08.04.2016. | ||
* Resuspension of oligo using the IDT protocole. | * Resuspension of oligo using the IDT protocole. | ||
* PCR of DesB and DesC (GC enhencer, Q5 master Mix and gel extract on the good strips ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR of DesB and DesC (GC enhencer, Q5 master Mix and gel extract on the good strips ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Colony PCR of CsgA, and gel electrophoresis ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR of CsgA, and gel electrophoresis ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | + | </div> | |
| − | + | ||
| − | + | ||
| − | + | ||
<div id="2016-08-09" class="evday"> | <div id="2016-08-09" class="evday"> | ||
| − | + | === Tuesday, August 9th === | |
* Colony PCR from transformation of the 08.08.2016 (size waited : 3000pb), clone1 has been sent to the sequencing. | * Colony PCR from transformation of the 08.08.2016 (size waited : 3000pb), clone1 has been sent to the sequencing. | ||
| Line 542: | Line 331: | ||
- 98°C 30s, (98°C 10s, 60°C 30s, 72°C 1min)*30, 72°C 2min. | - 98°C 30s, (98°C 10s, 60°C 30s, 72°C 1min)*30, 72°C 2min. | ||
- 98°C 30s, (98°C 10s, 72°C 1min)*30, 72°C 2min. | - 98°C 30s, (98°C 10s, 72°C 1min)*30, 72°C 2min. | ||
| − | + | </div> | |
| − | + | ||
<div id="2016-08-10" class="evday"> | <div id="2016-08-10" class="evday"> | ||
| − | + | === Wednesday, August 10th === | |
* Plasmid purification of Bba_K1951004 (248,49ng/µL) in pSB1K3 and sent to sequencing ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification of Bba_K1951004 (248,49ng/µL) in pSB1K3 and sent to sequencing ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| Line 552: | Line 340: | ||
* PCR clean up realised on PCR from the 08.09.2016, DesB (35ng/µL) and DesD (48ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up realised on PCR from the 08.09.2016, DesB (35ng/µL) and DesD (48ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
* SLIC realised from the previous PCR in pSB1C3 and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | * SLIC realised from the previous PCR in pSB1C3 and transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | ||
| − | + | </div> | |
| − | + | ||
<div id="2016-08-11" class="evday"> | <div id="2016-08-11" class="evday"> | ||
| − | + | === Thursday, August 11th === | |
* Sequencing of BBa_B0034+BBa_I0500 and Bba_K1951007 have been confirmed. | * Sequencing of BBa_B0034+BBa_I0500 and Bba_K1951007 have been confirmed. | ||
| Line 562: | Line 349: | ||
* Digestion of Bba_K1951007 by PstI and XbaI at 37°C during 1h ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion of Bba_K1951007 by PstI and XbaI at 37°C during 1h ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation over night of Bba_K1951007 and Bba_K880005 to create Bba_K1951010 (CsgA producer) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation over night of Bba_K1951007 and Bba_K880005 to create Bba_K1951010 (CsgA producer) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | + | </div> | |
| − | + | ||
<div id="2016-08-12" class="evday"> | <div id="2016-08-12" class="evday"> | ||
| − | + | === Friday, August 12th === | |
* Colony PRC from 08.10.2016 transformation again (DesB and DesD) and any clones were migrated to the good size again ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PRC from 08.10.2016 transformation again (DesB and DesD) and any clones were migrated to the good size again ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Plasmid purification on Bba_K1951009 (116.84ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification on Bba_K1951009 (116.84ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Ligation over night of Bba_K1951007 and Bba_K880005 to create Bba_K1951010 (CsgA producer) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Ligation_transformation Protocol#4]). | * Ligation over night of Bba_K1951007 and Bba_K880005 to create Bba_K1951010 (CsgA producer) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Ligation_transformation Protocol#4]). | ||
| − | + | </div> | |
| − | + | ||
<div id="2016-08-16" class="evday"> | <div id="2016-08-16" class="evday"> | ||
| − | + | === Tuesday, August 16th === | |
* Sequencing of Bba_K1951002 has been confirmed. | * Sequencing of Bba_K1951002 has been confirmed. | ||
| Line 581: | Line 366: | ||
* Colony PCR of Bba_K1951001 and Bba_K1951003 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR of Bba_K1951001 and Bba_K1951003 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Screening on agarose gel; 7 clones have been mixed in a PCR tube, and mix in 1µL used as DNA template. | * Screening on agarose gel; 7 clones have been mixed in a PCR tube, and mix in 1µL used as DNA template. | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-17" class="evday"> | <div id="2016-08-17" class="evday"> | ||
| − | + | === Wednesday, August 17th === | |
* Made petri dishes (10 50µG/mL kanamycin and 10 50µG/mL ampicillin). | * Made petri dishes (10 50µG/mL kanamycin and 10 50µG/mL ampicillin). | ||
| Line 591: | Line 375: | ||
* SLIC of DesB and DesD in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | * SLIC of DesB and DesD in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). | ||
* Transformation of the previous SLIC in compétante Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of the previous SLIC in compétante Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-18" class="evday"> | <div id="2016-08-18" class="evday"> | ||
| − | + | === Thursday, August 18th === | |
* Plasmid purification of Bba_K1951007(203.18ng/µL), Bba_K1951003(187.53ng/µL), Bba_K1951001 (188.74ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification of Bba_K1951007(203.18ng/µL), Bba_K1951003(187.53ng/µL), Bba_K1951001 (188.74ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| Line 602: | Line 385: | ||
* Digestion of Bba_K1951005, Bba_K1951006, Bba_K1951005 by XbaI and PstI ; Bba_K1951007 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion of Bba_K1951005, Bba_K1951006, Bba_K1951005 by XbaI and PstI ; Bba_K1951007 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Migration of the previous biobrick (size waited : Bba_K1951005=1547pb, Bba_K1951006=944pb, Bba_K1951007=506pb). | * Migration of the previous biobrick (size waited : Bba_K1951005=1547pb, Bba_K1951006=944pb, Bba_K1951007=506pb). | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-19" class="evday"> | <div id="2016-08-19" class="evday"> | ||
| − | + | === Friday, August 19th === | |
* Ligation of the digestion from the 08.18.2016 using 3A method in both pSB1A3 and pSB1C3 at room temperature during 1h ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of the digestion from the 08.18.2016 using 3A method in both pSB1A3 and pSB1C3 at room temperature during 1h ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| Line 612: | Line 394: | ||
* Colony PCR on the SLIC transformation of the 08.17.2016, nothing has worked ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR on the SLIC transformation of the 08.17.2016, nothing has worked ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Sequencing of the Bba_K1951004 is ok (desA producer). | * Sequencing of the Bba_K1951004 is ok (desA producer). | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-22" class="evday"> | <div id="2016-08-22" class="evday"> | ||
| − | + | === Monday, August 22th === | |
* PCR using Q5 master with new FliC coli sequence (substitute Bbs1 site) Bba_K1951005 and Bba_K1951001, every SLIC have been validated by electrophoresis ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR using Q5 master with new FliC coli sequence (substitute Bbs1 site) Bba_K1951005 and Bba_K1951001, every SLIC have been validated by electrophoresis ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* PCR clean up on the 2 previous SLIC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * PCR clean up on the 2 previous SLIC ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
| − | * SLIC and Transformation of Bba_K1951005 and Bba_K1951001 in | + | * SLIC and Transformation of Bba_K1951005 and Bba_K1951001 in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#.23Protocol_7_:_SLIC_.28sequence-_and_ligation-independent_cloning.29 Protocol#10]). |
* Starter of Bba_K1951004, Bba_K1951009 and Bba_K1951010 to make a protein production test. | * Starter of Bba_K1951004, Bba_K1951009 and Bba_K1951010 to make a protein production test. | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-23" class="evday"> | <div id="2016-08-23" class="evday"> | ||
| − | + | === Tuesday, August 23th === | |
* Colony PCR from transformation of the 08.25.2016 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR from transformation of the 08.25.2016 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| Line 633: | Line 413: | ||
* Colony PCR from Bba_K1951008, Bba_K1951009 | * Colony PCR from Bba_K1951008, Bba_K1951009 | ||
* Starters from Bba_K1951008 and Bba_K1951009 that seem ok. | * Starters from Bba_K1951008 and Bba_K1951009 that seem ok. | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-24" class="evday"> | <div id="2016-08-24" class="evday"> | ||
| − | + | === Wednesday, August 24th === | |
* Plasmid purification from Bba_K1951008, Bba_K1951009 and Bba_K1951001 and send to the sequencing with oligo FW ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification from Bba_K1951008, Bba_K1951009 and Bba_K1951001 and send to the sequencing with oligo FW ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Colony PCR from Bba_K1951005, Bba_K1951008, Bba_K1951009 and Bba_K1951010 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR from Bba_K1951005, Bba_K1951008, Bba_K1951009 and Bba_K1951010 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-25" class="evday"> | <div id="2016-08-25" class="evday"> | ||
| − | + | === Thursday, August 25th === | |
* Digestion of Bba_K1951002 and Bba_K1951003 by XbaI and PstI, Bba_B0034 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion of Bba_K1951002 and Bba_K1951003 by XbaI and PstI, Bba_B0034 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| Line 651: | Line 429: | ||
<div id="2016-08-25" class="evday"> | <div id="2016-08-25" class="evday"> | ||
| − | + | === Friday, August 26th === | |
* Sequencing Bba_K1951009, Bba_K1951001, Bba_K1951008 and Bba_K1951010 ok with the FW oligo only. | * Sequencing Bba_K1951009, Bba_K1951001, Bba_K1951008 and Bba_K1951010 ok with the FW oligo only. | ||
* Preparation of the HPLC solution : | * Preparation of the HPLC solution : | ||
| Line 674: | Line 452: | ||
* Pcr clean up on the digestion from the 08.25.2016 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | * Pcr clean up on the digestion from the 08.25.2016 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.237_:_PCR_clean-up_and_gel_extraction Protocol#7]). | ||
* Ligation using 2A method of Bba_K1951002 or Bba_K1951003 and Bba_B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation using 2A method of Bba_K1951002 or Bba_K1951003 and Bba_B0034 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Transformation of the previous ligation in | + | * Transformation of the previous ligation in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Digestion of 500ng in 50µL total mix of Bba_K1951002 and Bba_K1951005 by XbaI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion of 500ng in 50µL total mix of Bba_K1951002 and Bba_K1951005 by XbaI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) overnight at 16°C ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) overnight at 16°C ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) + BBa_I0500 (Prom) overnight at 16°C ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) + BBa_I0500 (Prom) overnight at 16°C ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-29" class="evday"> | <div id="2016-08-29" class="evday"> | ||
| − | + | === Monday, August 29th === | |
* Sequencing Bba_K1951001, Bba_K1951005, Bba_K1951009 and Bba_K1951010 ok with the RV oligo only so VALIDATE. | * Sequencing Bba_K1951001, Bba_K1951005, Bba_K1951009 and Bba_K1951010 ok with the RV oligo only so VALIDATE. | ||
| Line 688: | Line 465: | ||
* Ligation of Bba_K1951004 digested E/P in pSB1C3 at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951004 digested E/P in pSB1C3 at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
NB : we wanted to transform a cadA mutant from KEIO bank so we can't use this biobrick in pSB1K3. | NB : we wanted to transform a cadA mutant from KEIO bank so we can't use this biobrick in pSB1K3. | ||
| − | |||
</div> | </div> | ||
<div id="2016-08-30" class="evday"> | <div id="2016-08-30" class="evday"> | ||
| − | + | === Tuesday, August 30th === | |
| − | + | [[File:T--Aix-Marseille--PCR-30-06.jpg|left|thumb|Migration of FliC from E.coli, FliC from desulfo, DesB and DesC. The awaited size for the 8 DesD+RBS is 2150bp, for the FliC 25-54 it's 1900bp.]] | |
* Colony PCR of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Colony PCR of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) + BBa_I0500 (Prom) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR of Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) and Bba_B0034 (rbs) + BBa_I0500 (Prom) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | * Transformation of Bba_K1951004 (DesA with pSB1C3) in | + | * Transformation of Bba_K1951004 (DesA with pSB1C3) in competent Tg1 cells ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Ligation of Bba_K1951004 digested E/P in pSB1C3 at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951004 digested E/P in pSB1C3 at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
NB : we wanted to transform a cadA mutant from KEIO bank so we can't use this biobrick in pSB1K3. | NB : we wanted to transform a cadA mutant from KEIO bank so we can't use this biobrick in pSB1K3. | ||
* Starters from intermediate biobirck Bba_K1951001 (DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) and Bba_K1951000(DesA),Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom). | * Starters from intermediate biobirck Bba_K1951001 (DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) and Bba_K1951000(DesA),Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom). | ||
| − | * Made | + | * Made competent cells Tg1, culture from an over night starter have grown up from Abs(600nm)=0.05 until Abs(600nm)=0.457 at 37°C ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). |
</div> | </div> | ||
<div id="2016-09-01" class="evday"> | <div id="2016-09-01" class="evday"> | ||
| − | + | === Thursday, September 1st === | |
* Plasmid purification of the starter Bba_K1951001(175ng/µL), Bba_K1951002(250ng/µL), Bba_K1951003(440ng/µL) and Bba_K1951005(183ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification of the starter Bba_K1951001(175ng/µL), Bba_K1951002(250ng/µL), Bba_K1951003(440ng/µL) and Bba_K1951005(183ng/µL) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
| Line 711: | Line 487: | ||
* Ligation of Bba_K1951004 (S/P) and Bba_K1951001 + RBS (X/P) in pSB1K3(E/P) at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951004 (S/P) and Bba_K1951001 + RBS (X/P) in pSB1K3(E/P) at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation of Bba_K1951002 + RBS (E/S) and Bba_K1951003 + RBS (X/P) in pSB1K3(E/P) at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of Bba_K1951002 + RBS (E/S) and Bba_K1951003 + RBS (X/P) in pSB1K3(E/P) at 16°C overnight ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Made | + | * Made competent cells of fliC mutant and cadA mutant from the Keio bank and test on different antibiotic petri dishes (only kanamycin dish was a positive control) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). |
* Retest of colony PCR on Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) and nothing was good ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Retest of colony PCR on Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) and nothing was good ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | * Transformation of the ligation Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) from the ligation of the 08.26.2016 with the new | + | * Transformation of the ligation Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) from the ligation of the 08.26.2016 with the new competent cells from 08.30.2016 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
| − | * Transformation of Bba_K1951008 in fliC mutant Keio | + | * Transformation of Bba_K1951008 in fliC mutant Keio competent made the 30th, spread on Kana/Cm petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Made a calibration range for HPLC. | * Made a calibration range for HPLC. | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-09-02" class="evday"> | <div id="2016-09-02" class="evday"> | ||
| − | + | === Friday, September 2rd === | |
* Remade ligation of pSB1C3 (E/P) and Bba_K1951004 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Remade ligation of pSB1C3 (E/P) and Bba_K1951004 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Transformation of the previous ligation mix in | + | * Transformation of the previous ligation mix in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Colony PCR on transformation of intermediate biobricks ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]).: | * Colony PCR on transformation of intermediate biobricks ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]).: | ||
- Bba_K1951004 + RBS (X/P) Bba_K1951001 in pSB1K3(E/P) at 16°C overnight | - Bba_K1951004 + RBS (X/P) Bba_K1951001 in pSB1K3(E/P) at 16°C overnight | ||
- RBS Bba_K1951002(E/S) and RBS Bba_K1951003(X/P) in pSB1K3(E/P) at 16°C overnight | - RBS Bba_K1951002(E/S) and RBS Bba_K1951003(X/P) in pSB1K3(E/P) at 16°C overnight | ||
| − | |||
</div> | </div> | ||
<div id="2016-09-05" class="evday"> | <div id="2016-09-05" class="evday"> | ||
| − | + | === Monday, September 5th === | |
* Analysis of the electrophoresis results from 09.02.2016. | * Analysis of the electrophoresis results from 09.02.2016. | ||
| Line 742: | Line 515: | ||
<div id="2016-09-06" class="evday"> | <div id="2016-09-06" class="evday"> | ||
| − | + | === Tuesday, September 6th === | |
| − | + | [[File:T--Aix-Marseille--PCR-06-09.jpg|left|thumb|PCR colony verification. The holes 2 to 7 are the clone 12 of the plate and 11 to 16 are the clone 13 of the plate. The holes 9 and 17 are the RPF (control).]] | |
| − | * Transformation of cadA mutant | + | * Transformation of cadA mutant competent strain by Bba_K1951004 in pSB1C3, spread on Kana/Cm petri dishes ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Colony PCR Bba_K1951004 in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR Bba_K1951004 in pSB1C3 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* Starters from Bba_K1951004 in pSB1C3. | * Starters from Bba_K1951004 in pSB1C3. | ||
* 1 step of the transduction using phage p1 from fliC mutant Keio bank. Conserved at 4°C with 20µL chlorophorme | * 1 step of the transduction using phage p1 from fliC mutant Keio bank. Conserved at 4°C with 20µL chlorophorme | ||
* PCR with Q5 master mix using oligo of diriged mutagenesis for the insertion of Bbs1 site in the 214-215 and 238-239 sites of BBa_K1951008. Digest 3h with addition of 1µL DpNI in the PCR tube ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * PCR with Q5 master mix using oligo of diriged mutagenesis for the insertion of Bbs1 site in the 214-215 and 238-239 sites of BBa_K1951008. Digest 3h with addition of 1µL DpNI in the PCR tube ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | * Transformation of the previous PCR in | + | * Transformation of the previous PCR in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Starters of the PCR from 09.02.2016. | * Starters of the PCR from 09.02.2016. | ||
* Remade PCR on Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) but still no result ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Remade PCR on Bba_K1951001(DesB), Bba_K1951002(DesC), Bba_K1951003(DesD) with Bba_B0034 (rbs) + BBa_I0500(Prom) but still no result ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-09-07" class="evday"> | <div id="2016-09-07" class="evday"> | ||
| − | + | === Wednesday, September 7th === | |
* Colony PCR on cadA mutant complemented with BBa_K1951004 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR on cadA mutant complemented with BBa_K1951004 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
| Line 763: | Line 534: | ||
* Colony PCR to verify the insertion of Bbs1 site in the 214-215 and 238-239 sites of BBa_K1951008 and BBa_K1951009 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | * Colony PCR to verify the insertion of Bbs1 site in the 214-215 and 238-239 sites of BBa_K1951008 and BBa_K1951009 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). | ||
* From a starter of intermediate biobrick BBa_K1951004+ RBS BBa_K1951001, made culture at DO=0.2 until DO=0.6 and took a sample of 1uDO. Added 50µL beta mercatoethanol to the sample and congeled at -20°C to test protein production on a SDS page/Comassie. | * From a starter of intermediate biobrick BBa_K1951004+ RBS BBa_K1951001, made culture at DO=0.2 until DO=0.6 and took a sample of 1uDO. Added 50µL beta mercatoethanol to the sample and congeled at -20°C to test protein production on a SDS page/Comassie. | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-09-08" class="evday"> | <div id="2016-09-08" class="evday"> | ||
| − | + | === Thursday, September 8th === | |
* Starters of BBa_K1951008 + BbsI 215-216 position and starters of BBa_K1951008 + BbsI 238-239 position. | * Starters of BBa_K1951008 + BbsI 215-216 position and starters of BBa_K1951008 + BbsI 238-239 position. | ||
| Line 774: | Line 543: | ||
* Digestion of intermediate biobrick BBa_K1951004+ RBS BBa_K1951001 by EcoRI and SpeI and RBS BBa_K1951001+ RBS BBa_K1951002 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion of intermediate biobrick BBa_K1951004+ RBS BBa_K1951001 by EcoRI and SpeI and RBS BBa_K1951001+ RBS BBa_K1951002 by SpeI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Ligation of the 2 previous digestions in pSB1C3 to make the final BBa_K1951011 (E/P) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of the 2 previous digestions in pSB1C3 to make the final BBa_K1951011 (E/P) ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Transformation of the previous ligation in | + | * Transformation of the previous ligation in competent Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
| − | + | ||
</div> | </div> | ||
<div id="2016-09-09" class="evday"> | <div id="2016-09-09" class="evday"> | ||
| − | + | === Friday, September 9th === | |
| − | * Derivatization of the sample cadA mutant complemented with BBa_K1951004 and cadA mutant complemented with BBa_K1951004+ RBS BBa_K1951001 | + | * Derivatization of the sample cadA mutant complemented with BBa_K1951004 and cadA mutant complemented with BBa_K1951004+ RBS BBa_K1951001 . |
| − | * Reisolement of the knockout fliC mutant W3110 from the transformation of the 09.08.2016 | + | * Reisolement of the knockout fliC mutant W3110 from the transformation of the 09.08.2016. |
* Plasmid purification of the BBa_K1951008 + BbsI 215-216 position and starters of BBa_K1951008 + BbsI 238-239 position ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | * Plasmid purification of the BBa_K1951008 + BbsI 215-216 position and starters of BBa_K1951008 + BbsI 238-239 position ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Plasmid_DNA_purification Protocol#4]). | ||
* Digestion test by BBs1 and EcoRI using NEB buffer and electrophoresis analyse ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Digestion test by BBs1 and EcoRI using NEB buffer and electrophoresis analyse ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
| − | * Colony PCR of BBa_K1951011 in pSB1C3 by amplification with different SLIC oligos FW and RV to see the differente constitutive biobrick. Biobrick is OK. | + | * Colony PCR of BBa_K1951011 in pSB1C3 by amplification with different SLIC oligos FW and RV to see the differente constitutive biobrick. Biobrick is OK ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_PCR_and_electrophoresis Protocol#3]). |
| − | + | ||
| − | + | ||
</div> | </div> | ||
<div id="2016-09-10" class="evday"> | <div id="2016-09-10" class="evday"> | ||
| − | + | === Satursday, September 10th === | |
| − | * Swimming test of the knockout fliC mutant W3110 for 10 differents colonies and the W3110 WT as a positive temoin. | + | * Swimming test of the knockout fliC mutant W3110 for 10 differents colonies and the W3110 WT as a positive temoin ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.2311_:_Swimming_test Protocol#11]). |
* Starters of the fliC mutant W3110. | * Starters of the fliC mutant W3110. | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-09-11" class="evday"> | <div id="2016-09-11" class="evday"> | ||
| − | + | === Sunday, September 11st === | |
| − | * Made | + | * Made competent of the fliC mutant W3110 strain from 5 differents colonies from 4 colonies which swam well ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.231_:_Preparation_of_competent_bacteria_cells Protocol#1]). |
| − | * Transformation of the previous | + | * Transformation of the previous competent with BBa_K1951008 and BBa_K1951009 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
* Transformation of cadA mutant Keio strain with BBa_K1951011 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation of cadA mutant Keio strain with BBa_K1951011 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
<div id="2016-09-12" class="evday"> | <div id="2016-09-12" class="evday"> | ||
| − | + | === Monday, September 12rd === | |
| − | * Transformation of the fliC mutant W3110 | + | * Transformation of the fliC mutant W3110 competent with BBa_K1951008 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). |
| − | + | ||
</div> | </div> | ||
<div id="2016-09-13" class="evday"> | <div id="2016-09-13" class="evday"> | ||
| − | + | === Tuesday, September 13th === | |
| − | * Swimming test of colonies obtained after transformation of the 09.12.2016. After 3h incubation at 37°C, swimming has been recovered. | + | * Swimming test of colonies obtained after transformation of the 09.12.2016. After 3h incubation at 37°C, swimming has been recovered ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.2311_:_Swimming_test Protocol#11]). |
* Monitoring growth from differents strain to make comassie. From starters of fliC mutant W3110 BBa_K1951008, liC mutant W3110 BBa_K1951009, Tg1 BBa_K1951010, Tg1 WT (negative temoin), Tg1+pSB1C3 containing RFP (negative temoin), and cadA mutant BBa_K1951011, we made cultures at DO=0.2 in duplicate. At DO=0.6, half of the cultures has been induced with 0.02 arabinose for the biobrick containing an inductible promotor. Every hour, a sample of 1uDO was taken. After centrifugation at 5000g during 5min and removing of the supernatant, we added 50µL beta mercatoethanol and congeled at -20°C to test protein production on a SDS page/Comassie later. | * Monitoring growth from differents strain to make comassie. From starters of fliC mutant W3110 BBa_K1951008, liC mutant W3110 BBa_K1951009, Tg1 BBa_K1951010, Tg1 WT (negative temoin), Tg1+pSB1C3 containing RFP (negative temoin), and cadA mutant BBa_K1951011, we made cultures at DO=0.2 in duplicate. At DO=0.6, half of the cultures has been induced with 0.02 arabinose for the biobrick containing an inductible promotor. Every hour, a sample of 1uDO was taken. After centrifugation at 5000g during 5min and removing of the supernatant, we added 50µL beta mercatoethanol and congeled at -20°C to test protein production on a SDS page/Comassie later. | ||
| − | * HPLC test of the fliC mutant complemented with BBa_K1951011 | + | * HPLC test of the fliC mutant complemented with BBa_K1951011 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.239_:_Cadaverin_HPLC_analysis Protocol#9]). |
| − | + | ||
| − | + | ||
</div> | </div> | ||
<div id="2016-09-15" class="evday"> | <div id="2016-09-15" class="evday"> | ||
| − | + | === Thursday, September 15=== | |
* Dillution of peptides oligos by 10000 (ex: 0.31mg/238µL/10000=ng/µL). | * Dillution of peptides oligos by 10000 (ex: 0.31mg/238µL/10000=ng/µL). | ||
| Line 833: | Line 592: | ||
* Ligation of 5ng from the annealing mix for 100ng pSB1C3 digested by EcoRI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | * Ligation of 5ng from the annealing mix for 100ng pSB1C3 digested by EcoRI and PstI ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.233_:_Cloning_protocol_for_IDT_sequences Protocol#5]). | ||
* Transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | * Transformation in Tg1 ([https://2016.igem.org/Team:Aix-Marseille/Experiments/Protocols#Protocol_.232_:_Transformations Protocol#2]). | ||
| − | |||
| − | |||
</div> | </div> | ||
{{:Team:Aix-Marseille/Template-Footer}} | {{:Team:Aix-Marseille/Template-Footer}} | ||
